The only current effective treatment for obesity is surgical intervention. Novel treatments for obesity that avoid surgery are a desirable target. We investigate the mechanisms that attenuate the gut-brain vagal afferent pathway, resulting in loss of satiety signals from the gut that normally regulate food intake to provide peripheral targets fo novel therapies to effectively treat obesity. The ability of these gut-derived satiation signals to activate vagal afferent neurons and decrease food intake is absent in rodent models of diet-induced obesity (DIO), and altered gut-brain signaling has been shown to be present in human obesity. We have shown that an early response in high fat (HF) DIO is the onset of leptin resistance in vagal afferent neurons. To show causality, we developed a novel mouse model with a selective deletion of the leptin receptor in vagal afferent neurons. These mice gain weight and have increased adiposity on a regular chow diet, and do not increase their weight further on a high fat diet as would be expected. Thus, impaired leptin signaling in vagal afferent neurons is sufficient and necessary for an obese phenotype. We will now address the mechanisms that lead to leptin resistance in vagal afferent neurons, whether it can be reversed and if so, will thi reverse DIO. Obesity in humans and in rodent models is associated with impaired intestinal barrier function and metabolic endotoxemia, defined as low, chronic increases in plasma levels of the bacterial product lipopolysaccharide (LPS). In vagal afferent neurons, leptin-resistance can be induced by chronic treatment with low dose LPS and is accompanied by Toll-like receptor 4 (TLR4; receptor for LPS) activation and its downstream signal MyD88. Exposure of vagal afferent neurons to LPS in vitro increases SOCS3 and decreases leptin-induced pSTAT3. In the proposed research we will test the hypothesis that manipulation of the gut microbiota can restore leptin sensitivity in vagal afferent neurons and reverse DIO. The hypothesis will be addressed in two specific aims. In AIM 1, we will determine the intracellular signaling pathways leading to reversal of leptin resistance in vagal afferent neurons. We expect to find that: 1. leptn resistance in vagal afferent neurons is dependent on TLR4-MyD88 and/or Ghrelin receptor elevation of SOCS3, and 2. silencing SOCS3 or MyD88 will restore leptin sensitivity and reverse DIO. In AIM 2, expect to show that reversing the changes in intestinal barrier function using GLP2 which restores barrier function and normalizing gut microbiota using a inulin as a prebiotic can restore leptin sensitivity in vagal afferent neurons and reverses DIO. It is critical to understand the mechanisms by which vagal afferent neurons develop leptin resistance and how this can be reversed to restore satiety signaling from the gut. This proposal will take the novel approach of normalizing gut microbiota and barrier function to reverse leptin resistance. Defining both leptin sensitivity and changes in gut microbiota and barrier function are both priorities as the gut and vagal afferent neurons are peripheral targets for therapeutic interventio to solve the pressing need to treat obesity.